Example structure of a “good” guide
## Warning: Removed 1 rows containing non-finite values (stat_ydensity).
## [1] "mixed model failed: NCR_1411"
Per guide: * Per timepoint: t-test for difference in signal between no activator and 100 fM conditions per timepoint * Perform FDR correction for number of measurements from start of experiment to timepoint * Return first timepoint for which corrected p-value < 0.05
Structures of the two guides that performed well (rate > 1 above background) but without hairpin structure (NCR_1346, NCR_1351):
Determination of how much of predicted hairpin structure needs to be maintained:
## NCR.id spacer
## 120 NCR_1313 GUUUACCUUGGUAAUCAUCU
## 126 NCR_1319 UCAUUAAAUGGUAGGACAGG
## 137 NCR_1330 GCAAUCAAUGGGCAAGCUUU
## 138 NCR_1331 CUUCUCUGUAGCUAGUUGUA
## 139 NCR_1332 GAGUAAAUCUUCAUAAUUAG
## 142 NCR_1335 AUGGUGUCCAGCAAUACGAA
## 143 NCR_1336 GCCGUCUUUGUUAGCACCAU
## 155 NCR_1348 AUUAGCUCUCAGGUUGUCUA
## 156 NCR_1349 UGGUACGUUAAAAGUUGAUG
## 158 NCR_1351 UGGCUACUUUGAUACAAGGU
## 21685 NCR_1410 UGAAUGUAAAACUGAGGAUCUGAAAACU
## 9671 NCR_1412 UAUAAGCAAUUGUUAUCCAGAAAGGUAC
## 10691 NCR_1417 GAUUGAGAAACCACCUGUCUCCAUUUAU
## structure
## 120 ...((((((((.........))))................))))........
## 126 ...((((((((.........))))................))))........
## 137 .......(((((....(((...((........))..))).))))).......
## 138 ..(((.(((........(((((.........)))))......))).)))...
## 139 ...............(((((((..(((......)))...)))))))......
## 142 ................((..((((((((.....))))))))..)).......
## 143 ..............((((((((((........)))..)))))))........
## 155 (((((.((((......(((.((((............))))))))))))))))
## 156 ...((((((((.........))))........)))).((((((...))))))
## 158 (((.(((((((.........))))........))))))(((((...))))).
## 21685 ((((((.((((.........))))......((.....))........)).))))......
## 9671 ...(((.((((.........))))............(((....))).........)))..
## 10691 ...............(((((((((((......................)))))).)))))
## Warning in cor.test.default(GC_content, Estimate, method = "spearman"): Cannot
## compute exact p-value with ties
## Warning in cor.test.default(GC_content, Estimate, method = "spearman"): Cannot
## compute exact p-value with ties
## Warning in is.na(x): is.na() applied to non-(list or vector) of type 'language'
## Warning in is.na(x): is.na() applied to non-(list or vector) of type 'language'
## Warning in cor.test.default(downstream_U, Estimate, method = "spearman"): Cannot
## compute exact p-value with ties
## Warning in cor.test.default(downstream_U, Estimate, method = "spearman"): Cannot
## compute exact p-value with ties
## Warning in cor.test.default(downstream_unstructured_u, Estimate, method =
## "spearman"): Cannot compute exact p-value with ties
## Warning in cor.test.default(downstream_unstructured_u, Estimate, method =
## "spearman"): Cannot compute exact p-value with ties
## Warning in is.na(x): is.na() applied to non-(list or vector) of type 'language'
## Warning in is.na(x): is.na() applied to non-(list or vector) of type 'language'
## Warning in is.na(x): is.na() applied to non-(list or vector) of type 'language'
## Warning in is.na(x): is.na() applied to non-(list or vector) of type 'language'
## Warning in predict.lm(training_fit, newdata = subset(lan_regression_data, :
## prediction from a rank-deficient fit may be misleading
## Likelihood ratio test
##
## Model 1: rate ~ spacer_1_A + spacer_1_C + spacer_1_G + spacer_1_U + spacer_2_A +
## spacer_2_C + spacer_2_G + spacer_2_U + spacer_3_A + spacer_3_C +
## spacer_3_G + spacer_3_U + spacer_4_A + spacer_4_C + spacer_4_G +
## spacer_4_U + spacer_5_A + spacer_5_C + spacer_5_G + spacer_5_U +
## spacer_6_A + spacer_6_C + spacer_6_G + spacer_6_U + spacer_7_A +
## spacer_7_C + spacer_7_G + spacer_7_U + spacer_8_A + spacer_8_C +
## spacer_8_G + spacer_8_U + spacer_9_A + spacer_9_C + spacer_9_G +
## spacer_9_U + spacer_10_A + spacer_10_C + spacer_10_G + spacer_10_U +
## spacer_11_A + spacer_11_C + spacer_11_G + spacer_11_U + spacer_12_A +
## spacer_12_C + spacer_12_G + spacer_12_U + spacer_13_A + spacer_13_C +
## spacer_13_G + spacer_13_U + spacer_14_A + spacer_14_C + spacer_14_G +
## spacer_14_U + spacer_15_A + spacer_15_C + spacer_15_G + spacer_15_U +
## spacer_16_A + spacer_16_C + spacer_16_G + spacer_16_U + spacer_17_A +
## spacer_17_C + spacer_17_G + spacer_17_U + spacer_18_A + spacer_18_C +
## spacer_18_G + spacer_18_U + spacer_19_A + spacer_19_C + spacer_19_G +
## spacer_19_U + spacer_20_A + spacer_20_C + spacer_20_G + spacer_20_U
## Model 2: rate ~ spacer_1_A + spacer_1_C + spacer_1_G + spacer_1_U + spacer_2_A +
## spacer_2_C + spacer_2_G + spacer_2_U + spacer_3_A + spacer_3_C +
## spacer_3_G + spacer_3_U + spacer_4_A + spacer_4_C + spacer_4_G +
## spacer_4_U + spacer_5_A + spacer_5_C + spacer_5_G + spacer_5_U +
## spacer_6_A + spacer_6_C + spacer_6_G + spacer_6_U + spacer_7_A +
## spacer_7_C + spacer_7_G + spacer_7_U + spacer_8_A + spacer_8_C +
## spacer_8_G + spacer_8_U + spacer_9_A + spacer_9_C + spacer_9_G +
## spacer_9_U + spacer_10_A + spacer_10_C + spacer_10_G + spacer_10_U +
## spacer_11_A + spacer_11_C + spacer_11_G + spacer_11_U + spacer_12_A +
## spacer_12_C + spacer_12_G + spacer_12_U + spacer_13_A + spacer_13_C +
## spacer_13_G + spacer_13_U + spacer_14_A + spacer_14_C + spacer_14_G +
## spacer_14_U + spacer_15_A + spacer_15_C + spacer_15_G + spacer_15_U +
## spacer_16_A + spacer_16_C + spacer_16_G + spacer_16_U + spacer_17_A +
## spacer_17_C + spacer_17_G + spacer_17_U + spacer_18_A + spacer_18_C +
## spacer_18_G + spacer_18_U + spacer_19_A + spacer_19_C + spacer_19_G +
## spacer_19_U + spacer_20_A + spacer_20_C + spacer_20_G + spacer_20_U +
## structure_1_. + structure_1_structured + structure_1_unstructured +
## structure_2_. + structure_2_both + structure_2_structured +
## structure_2_unstructured + structure_3_. + structure_3_both +
## structure_3_structured + structure_3_unstructured + structure_4_. +
## structure_4_both + structure_4_structured + structure_4_unstructured +
## structure_5_. + structure_5_both + structure_5_structured +
## structure_5_unstructured + structure_6_. + structure_6_both +
## structure_6_structured + structure_6_unstructured + structure_7_. +
## structure_7_both + structure_7_structured + structure_7_unstructured +
## structure_8_. + structure_8_both + structure_8_structured +
## structure_8_unstructured + structure_9_. + structure_9_both +
## structure_9_structured + structure_9_unstructured + structure_10_. +
## structure_10_both + structure_10_structured + structure_10_unstructured +
## structure_11_. + structure_11_both + structure_11_structured +
## structure_11_unstructured + structure_12_. + structure_12_both +
## structure_12_structured + structure_12_unstructured + structure_13_. +
## structure_13_both + structure_13_structured + structure_13_unstructured +
## structure_14_. + structure_14_both + structure_14_structured +
## structure_14_unstructured + structure_15_. + structure_15_both +
## structure_15_structured + structure_15_unstructured + structure_16_. +
## structure_16_both + structure_16_structured + structure_16_unstructured +
## structure_17_. + structure_17_both + structure_17_structured +
## structure_17_unstructured + structure_18_. + structure_18_both +
## structure_18_structured + structure_18_unstructured + structure_19_. +
## structure_19_both + structure_19_structured + structure_19_unstructured +
## structure_20_. + structure_20_both + structure_20_structured +
## structure_20_unstructured
## #Df LogLik Df Chisq Pr(>Chisq)
## 1 62 -799.34
## 2 106 -763.56 44 71.567 0.005376 **
## ---
## Signif. codes: 0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1
## Warning: glm.fit: fitted probabilities numerically 0 or 1 occurred
Model 1: sequence + structure
Model 2: only sequence
Model 3: only structure
Model 4: only sequence (binary)
Model 5: sequence (binary) + structure
Top guides:
Bottom guides:
## Warning in eval(substitute(expr), data, enclos = parent.frame()): NAs introduced
## by coercion
## Warning: NAs introduced by coercion
## [1] "mixed model failed: NCR_1320"
## [1] "mixed model failed: NCR_1332"
## [1] "mixed model failed: NCR_1387"
## Warning: Removed 27 rows containing non-finite values (stat_smooth).
## Warning: Removed 27 rows containing missing values (geom_point).
## Warning: Removed 5 rows containing missing values (geom_smooth).
gBlock round 2 outlier: